Also the modification in compound 6, inspired by albomycin whereby pepN hydrolyzes the peptide bond between a serine and a modified amino acid carrying an acidic side chain, could not rescue the activity. From the in vitro aminoacylation experiments it can also be concluded that lack of whole cell activity is due to inability of the peptidases to metabolize peptides containing -amino acids. Hence, this shows that the peptidases PepA, PepB and PepN, commonly known to be responsible for processing of McC and its analogues, only can cleave these compounds as exopeptidases and are not able to release the GDC-0449 supply active moiety via endopeptidic cleavage. Whether or not incorporation of -amino acids in the transport peptide part RAD001 interferes with uptake of the McC derivatives by the YejABEF transporter is less relevant here, as in vitro tests already show lack of activity. Additional reduced uptake of the analogues 4-7 however cannot be excluded. The observation that -D-SA and its McC derivative 8 can inhibit AspRS, shows that the peptidases can metabolize peptide bonds between two amino acids, whereby the Cterminal amino acid has the -configuration -Asp). This suggests that the peptidases involved in this reaction are only stereoselective for the N-terminal amino acid. Since it was already frequently observed that -Asp can be esterified to tRNA, the finding that -D-SA can also inhibit AspRS can be considered being an expected result. This also shows that the absolute configuration of the amino acid is not required for recognition inside the active site of AspRS, and most probably the same holds for other aaRSs. This finding matches with the results of Thompson et al. who concluded that there is only limited chiral specificity for L-Asp, leading to an esterification of -Asp to tRNAAsp with a rate of 1:4000 for -Asp vs -Asp. These results are conflicting however with the views of Banik and Nandi who studied the chiral discrimination by enzymes in protein synthesis via semi-empirical calculation methods. From their theoretical studies they concluded that the network of electrostatic interactions between the incoming amino acid, ATP and the synthetase are highly unfavorable for incorporation of a -amino acid. Not only in the aminoacylation step, but likewise in the peptide bond formation reactions, it would be virtually impossible to incorporate -amino acids in protein structures. Our results however clearly demonstrate the in vitro inhibitory effects and hence recognition of our -amino acid containing aminoacyl adenylate analogues.